The invention relates to the synthesis of arrays of chemical compounds. A variety of chemical compounds can be synthesized on a solid support. Combinatorial libraries of chemical compounds can be synthesized on the solid phase (see, e.g., DeWitt et al. (1993) Proc Nat Acad USA 90:6909; and Bunin et al. (1994) 91:4708). For example, Merrifield developed a method for synthesizing peptides on a support (Merrifield (1965) Science 150:178-185). This method was extended to oligonucleotides by Letsinger and others (see, e.g., Beaucage and Iyer (1992) Tetrahedron 12:2223 for a review). Arrays of chemical compounds can be produced by synthesizing the different compounds on a common solid support, e.g., a planar surface of the support. Such arrays provide a convenient format for analyzing multiple interactions in parallel, and, accordingly, is adaptable for high throughput screening, e.g., for drug discovery, gene discovery, genotyping, and diagnosis.
Arrays can be produced by the in situ synthesis of compounds on the solid support to form the covalent immobilization of compounds are in-situ synthesis of biopolymers from properly protected monomers.
Photolithographic methods (see, e.g., Fodor et al. (1991) Science 251:767-773; Fodor et al. (1993) Nature 364:555-556; U.S. Pat. No. 5,143,854) have been used to construct oligonucleotide arrays with spatial addressability. This approach, at least in some implementations, uses customized photomasks to control the addition of a particular nucleotide monomer at each successive addition reaction. In another approach, micromirrors (see, e.g., Gao et al. (2000) J. Comb. Chem. 2:349-354; M. R. Sussman et al. (1999) Nature Biotechnology 17:974-978; and WO 99/41007) are employed to provide an addressable array of light without the use of a photomask. The light can locally produce acid which subsequently removes the acid labile protecting groups, 4,4′-dimethoxy-trityl (DMT), on nucleotide monomers.
A third approach uses a pulse jet, such as found in an inkjet printer, to distribute sub-nanoliter volume of nucleotide monomer solution and the activation agent such as tetrazole (see, e.g., WO 95/25116, U.S. Pat. Nos. 6,028,189; 5,874,554; 5,474,796; 6,177,558 and WO 98/41531). The piezoelectric pumps deliver, e.g., 5′-protected, 3′-phosphoramidite activated nucleotides dissolved in organic solvent to individual spots on an array. The solution of activated nucleotide is mixed with a solution of a coupling agent to covalently attach the activated nucleotide to a functional group on the glass surface. The 5′-protecting groups are subsequently removed with a deprotecting reagent. The entire process is repeated until the desired oligonucleotides are formed.
In one implementation, five nozzle heads are used for DNA synthesis using the pulse jet method. Four nozzle heads are provided for the four nucleotide monomers A, T, G, C, and a fifth nozzle head is used to deliver a solution of an activating agent for coupling reactions after each nucleotide monomer is printed on the substrate. The accuracy of delivering the activating agent to a location where one of the nucleotide monomers is previously spotted can impact the coupling reaction (see, e.g., U.S. Pat. Nos. 5,985,551 and 5,474,796). The entire apparatus is kept in a moisture-free environment at every step in the process.